Research Paper Volume 16, Issue 10 pp 9251—9263

Figure 4. Inhibition of the NF-κB signaling pathway with PDTC reversed the effect of TMAO on osteoclast differentiation. (A, B) RAW 264.7 cells were pretreated with 25 μM PDTC for 2 h before being cultured with or without 100 μM TMAO. The number of TRAP-positive osteoclasts increased under 100 μM TMAO and decreased with PDTC pretreatment with or without TMAO. (magnification ×40). (C) The expression of β-CTx was up-regulated under 100 μM TMAO and suppressed by PDTC with or without TMAO treatment evaluated by ELISA (n = 3 per group). (D) The expression of osteoclast-specific genes was up-regulated under 100 μM TMAO and suppressed by PDTC with or without TMAO treatment evaluated by real-time PCR (n = 4 per group). (E, F) The protein expression levels of NFATc1, c-Fos, Cathepsin K, NF-κB p65, and Traf6 were promoted under 100 μM TMAO and suppressed by PDTC with or without TMAO treatment as examined by Western blot (n = 3 per group). * p < 0.05, ** p < 0.01.